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Reconstructed cell fate-regulatory programs in stem cells reveal hierarchies and key factors of neurogenesis.

ABSTRACT: Cell lineages, which shape the body architecture and specify cell functions, derive from the integration of a plethora of cell intrinsic and extrinsic signals. These signals trigger a multiplicity of decisions at several levels to modulate the activity of dynamic gene regulatory networks (GRNs), which ensure both general and cell-specific functions within a given lineage, thereby establishing cell fates. Significant knowledge about these events and the involved key drivers comes from homogeneous cell differentiation models. Even a single chemical trigger, such as the morphogen all-trans retinoic acid (RA), can induce the complex network of gene-regulatory decisions that matures a stem/precursor cell to a particular step within a given lineage. Here we have dissected the GRNs involved in the RA-induced neuronal or endodermal cell fate specification by integrating dynamic RXRA binding, chromatin accessibility, epigenetic promoter epigenetic status, and the transcriptional activity inferred from RNA polymerase II mapping and transcription profiling. Our data reveal how RA induces a network of transcription factors (TFs), which direct the temporal organization of cognate GRNs, thereby driving neuronal/endodermal cell fate specification. Modeling signal transduction propagation using the reconstructed GRNs indicated critical TFs for neuronal cell fate specification, which were confirmed by CRISPR/Cas9-mediated genome editing. Overall, this study demonstrates that a systems view of cell fate specification combined with computational signal transduction models provides the necessary insight in cellular plasticity for cell fate engineering. The present integrated approach can be used to monitor the in vitro capacity of (engineered) cells/tissues to establish cell lineages for regenerative medicine.

SUBMITTER: Mendoza-Parra MA

PROVIDER: S-EPMC5088593 | biostudies-literature | 2016 Nov

REPOSITORIES: biostudies-literature

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Reconstructed cell fate-regulatory programs in stem cells reveal hierarchies and key factors of neurogenesis.

Mendoza-Parra Marco-Antonio MA Malysheva Valeriya V Mohamed Saleem Mohamed Ashick MA Lieb Michele M Godel Aurelie A Gronemeyer Hinrich H

Genome research 20160920 11

Cell lineages, which shape the body architecture and specify cell functions, derive from the integration of a plethora of cell intrinsic and extrinsic signals. These signals trigger a multiplicity of decisions at several levels to modulate the activity of dynamic gene regulatory networks (GRNs), which ensure both general and cell-specific functions within a given lineage, thereby establishing cell fates. Significant knowledge about these events and the involved key drivers comes from homogeneous ...[more]

PMID: 27650846

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Project description:BackgroundDisruptions of SETBP1 (SET binding protein 1) on 18q12.3 by heterozygous gene deletion or loss-of-function variants cause SETBP1 disorder. Clinical features are frequently associated with moderate to severe intellectual disability, autistic traits and speech and motor delays. Despite the association of SETBP1 with neurodevelopmental disorders, little is known about its role in brain development.MethodsUsing CRISPR/Cas9 genome editing technology, we generated a SETBP1 deletion model in human embryonic stem cells (hESCs) and examined the effects of SETBP1-deficiency in neural progenitors (NPCs) and neurons derived from these stem cells using a battery of cellular assays, genome-wide transcriptomic profiling and drug-based phenotypic rescue.ResultsNeural induction occurred efficiently in all SETBP1 deletion models as indicated by uniform transition into neural rosettes. However, SETBP1-deficient NPCs exhibited an extended proliferative window and a decrease in neurogenesis coupled with a deficiency in their ability to acquire ventral forebrain fate. Genome-wide transcriptome profiling and protein biochemical analysis revealed enhanced activation of Wnt/β-catenin signaling in SETBP1 deleted cells. Crucially, treatment of the SETBP1-deficient NPCs with a small molecule Wnt inhibitor XAV939 restored hyper canonical β-catenin activity and restored both cortical and MGE neuronal differentiation.LimitationsThe current study is based on analysis of isogenic hESC lines with genome-edited SETBP1 deletion and further studies would benefit from the use of patient-derived iPSC lines that may harbor additional genetic risk that aggravate brain pathology of SETBP1 disorder.ConclusionsWe identified an important role for SETBP1 in controlling forebrain progenitor expansion and neurogenic differentiation. Our study establishes a novel regulatory link between SETBP1 and Wnt/β-catenin signaling during human cortical neurogenesis and provides mechanistic insights into structural abnormalities and potential therapeutic avenues for SETBP1 disorder.

Project description:Cell cycle proteins are important regulators of diverse cell fate decisions, and in this capacity have pivotal roles in neurogenesis and brain development. The mechanisms by which cell cycle regulation is integrated with cell fate control in the brain and other tissues are poorly understood, and an outstanding question is whether the cell cycle machinery regulates fate decisions directly or instead as a secondary consequence of proliferative control. Identification of the genes targeted by E2 promoter binding factor (E2f) transcription factors, effectors of the pRb/E2f cell cycle pathway, will provide essential insights into these mechanisms. We identified the promoter regions bound by three neurogenic E2f factors in neural precursor cells in a genome-wide manner. Through bioinformatic analyses and integration of published genomic data sets we uncovered hundreds of transcriptionally active E2f-bound promoters corresponding to genes that control cell fate processes, including key transcriptional regulators and members of the Notch, fibroblast growth factor, Wnt and Tgf-β signaling pathways. We also demonstrate a striking enrichment of the CCCTC binding factor transcription factor (Ctcf) at E2f3-bound nervous system-related genes, suggesting a potential regulatory co-factor for E2f3 in controlling differentiation. Finally, we provide the first demonstration of extensive tissue specificity among E2f target genes in mammalian cells, whereby E2f3 promoter binding is well conserved between neural and muscle precursors at genes associated with cell cycle processes, but is tissue-specific at differentiation-associated genes. Our findings implicate the cell cycle pathway as a widespread regulator of cell fate genes, and suggest that E2f3 proteins control cell type-specific differentiation programs by regulating unique sets of target genes. This work significantly enhances our understanding of how the cell cycle machinery impacts cell fate and differentiation, and will importantly drive further discovery regarding the mechanisms of cell fate control and transcriptional regulation in the brain, as well as in other tissues.

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Reconstructed cell fate-regulatory programs in stem cells reveal hierarchies and key factors of neurogenesis.

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Reconstructed cell fate-regulatory programs in stem cells reveal hierarchies and key factors of neurogenesis.

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